Project Summary Type 1 diabetes (T1D) results in the immune-mediated destruction of insulin-producing beta cells in the pancreas. As such, immune intervention to prevent, delay, or reverse T1D is an appealing therapeutic approach. Development of an antigen-specific intervention that targets the diabetogenic immune response without compromising systemic immunity is complicated by an expanding list of antigenic targets in T1D. Recently, novel, post-translationally modified hybrid fusion peptides generated at the site of autoimmune attack in the islet have also been identified as potent autoantigens. Thus, development of effective antigen-based therapy requires not only antigen identification, but also an understanding the unique antigen processing environment facilitated by the islet itself. This proposal seeks to identify the effects of lymphocyte metabolism on T-B interactions in T1D. Hypoxia is a potent regulator of B cell proliferation and expansion at the site of T-B interactions in the germinal center of primary lymphoid organs. Germinal centers also form in the islets during autoimmune attack; however, the effect of hypoxia on islet germinal centers is not known. These germinal centers are formed within areas of lymphocytic infiltrate, termed insulitis. Interestingly, not all insulitis lesions progress to beta cell death. Given the highly vascularized nature of the islet needed to accommodate the metabolic demand associated with insulin secretion, it is possible that the islet?s distinct metabolic microenvironment influences T and B cell interactions at the site of autoimmune attack in the pancreas. The proposed research will test the hypothesis that the sites of T-B cell interactions in the islet are metabolically distinct from those of primary lymphoid organs and that the metabolic profiles of infiltrating lymphocytes are differentially regulated to develop autoaggressive or regulatory phenotypes that either drive or protect against the development of T1D. In aim 1, we will assess how the metabolic environment in the islet shapes the immune effector response as diabetes progresses. A unique hypoxia probe will be used to assess changes in islet oxygen tension and a combination of metabolic phenotyping and real-time bioenergetic analysis will be used to identify changes in cellular energy sources throughout the disease process. In aim 2, transgenic NOD mouse models will be used to delineate the metabolic parameters that distinguish innocuous from destructive insulitis. These studies will identify characteristic metabolic features of autoimmunity and tolerance in T1D -- differences that may be harnessed for future therapeutic interventions.